Human serum albumin is the most abundant plasma protein of adults. The concentration of albumin is 40 mg/ml, or 160 g of albumin circulating throughout the human body for a 70 Kg adult male. This protein maintains osmotic pressure and functions in the binding and transport of copper, nickel, calcium (weakly, at 2-3 binding sites), bilirubin and protoporphyrin, long-chain fatty acids, prostaglandins, steroid hormones (weak binding with these hormones promotes their transfer across the membranes), thyroxine, triiodothyronine, crystine, and glutathione. According to Peters, T. and Reed, R. G. in Albumin: Structure, Biosynthesis and Function, (Peter, T. and Sjoholm, J. eds.) 1977 p. 11-20, over 10,000 kilograms of purified albumin are administered annually in the United States alone to patients with circulatory failure or with albumin depletion.
Currently the only commercial source of HSA is from fractionated blood. Considering the possible dangers of blood borne contaminants and pathogens, it would be a considerable contribution to the commercial production of HSA to develop alternate methods of producing HSA. With the advent of recombinant DNA technology, it is now possible to produce HSA by alternate methods.
HSA has also been expressed in Saccharomyces cerevisiae as disclosed by Etcheverry et al. in Bio/technology, August 1986, p. 726 and Arjum Singh in EPA 123,544. Etcheverry disclosed HSA expression intracellularly in a concentration of approximately 6 mg/l and the secretion of HSA which remained cell associated. Arjum Singh also disclosed the expression of HSA in Saccharomyces cerevisiae in combination with the .alpha.-factor promoter and signal sequence. Singh appears to have been able to achieve an intracellular production level of approximately 25 mg/l and a secreted production level of 3 mg/l. Pichia pastoris has also been used to express HSA as is disclosed in EPA 344,459. The concentration of HSA produced in Pichia pastoris appears to be about 89 ng HSA/mg of protein. Although the process for producing HSA in recombinant expression system has been established by these experiments it would be desirable to optimize these processes to achieve the maximum possible HSA production.
Therefore, it would be a significant contribution to the art to provide a process for increasing the yeild of HSA from the recombinant expression of HSA in microorganism such as Pichia pastoris.
Therefore, it is an object of this invention to provide a process for increasing the yield of HSA produced in a recombinant expression systems.